The mechanisms that lead to motor neuron dysfunction and motor neuron death in amyotrophic lateral sclerosis (ALS) are yet to be completely understood. Since excitability studies of acutely dissociated, viable spinal cord cells cannot be done in ALS patients, we propose to study excitability in viable, acutely dissociated spinal cord cell subpopulations of the Wobbler mouse, an autosomal recessive animal model for the study of motor neuron disease (MND). The goal of this study is the identification of abnormalities of neurotransmitter-receptor function in populations of viable Wobbler mouse cervical spinal cord cells. The rationale is that genetic factors are important determinants of various forms of human MND but the functional expression of the genetic defect is unknown. Since excitatory amino acids and related substances play a pathogenetic role in motor neuron cytotoxicity, the primary cellular defect might be an alteration of excitatory amino acid neurotransmitter-receptor function. The hypothesis of this project proposes that genetic abnormalities in the Wobbler mouse may be phenotypically expressed by abnormal responses to excitatory amino acid neurotransmitters in cervical spinal cord cells, those that are affected the most in the Wobbler. Electrical and chemical excitability will be studied in large populations of viable spinal cord cells using flow cytometry and voltage-sensitive dyes. Relative membrane potential changes to excitatory amino acids as well as to voltage-dependent neurotoxins, inhibitory amino acids and neuropeptides will be analyzed. Wobbler (NFR/wr) and control (NFR) spinal cords will be dissected in 6 regions and cells will be isolated using a papain dissociation protocol. Tracer studies, immunocytochemistry and gradient ultracentrifugation will permit membrane potential determinations in identifiable motor neurons and in other cell phenotypes. Data will be analyzed using the computers of the University of New Mexico and the Los Alamos National Laboratory. Single- cell quantitative fluorescence microscopy using voltage-sensitive dyes will complement the flow cytometer population data. A disability score analysis using clinical parameters will be used to correlate neuromuscular dysfunction with cellular abnormalities. The characterization of possible membrane potential abnormalities in response to amino acid neurotransmitters in identifiable subpopulations of Wobbler mouse spinal cord cells will provide unique information relevant to the pathogenesis of ALS.